Massive parallel single-nucleus transcriptomic analysis of post-infarct adult hearts

Abstract

Heart cell homeostasis changes significantly in diseases such as heart failure after myocardial infarction (MI). Due to the heterogeneity of cell populations in adult mammalian heart, it remained challenging to dissect the transcriptomic networks simultaneously for all cardiac cell types. While single-cell transcriptomic technologies are powerful in dealing with cells of similar morphology and phenotypes, they are not tailored to handle complex populations composed of both exceptionally large cells (e.g., adult cardiomyocytes, ACMs) and usual small cells (e.g., non-cardiomyocytes). Massive parallel single-nucleus RNA sequencing (snRNA-seq) method avoids the needs of isolating intact single cells or enriching specific cell populations, therefore it is well suits for analyzing highly complex tissues including mammalian hearts. Based on our previous finding that ACMs can dedifferentiate – becoming more primitive, and proliferate in vitro, we have since developed a transgenic triple-reporter mouse model to map ACMs and follow their dedifferentiation and proliferation, allowing the visualization of all 4 plausible modes of ACM regeneration in vivo. Using snRNA-seq, we obtained large datasets for both normal and post-MI hearts from our tri-transgenic mice, and identified major heart cell populations with significant heterogeneity of CMs, fibroblasts, endothelial cells, vascular smooth muscle cells, and macrophages. We compared dedifferentiated CMs with normal CMs based on their expressions of CM lineage marker and dedifferentiation reporter transgenes. We found that dedifferentiated CMs in post-MI hearts had significant down-regulations of gene networks and pathways for cardiac hypertrophy and metabolism, and contractile and rhythmic functions, but up-regulation in signaling pathways and gene sets for cell survival, active cell cycle and proliferation. Our data present an important resource and strategy for studying cardiovascular biology and provide insights into post-infarct cardiac remodeling using methodology readily applicable to other biomedical fields.